Among neurons that contain gonadotropin releasing hormone (GnRH) only a select sub-population, widely scattered in the forebrain, send their axons to the median eminence (ME). Previous studies of the distribution and synaptic connectivity of these cells have shown that the organization of the GnRH network undergoes systematic reordering during the lifetime of the individual. This proposal will extend those observations with the goal of identifying the neuronal and/or hormonal inputs to GnRH cells responsible for their physiological modulation. Four possible mechanisms of regulation will be studied. (1) Because the physical relationship between GnRH neurosecretory axons and the capillaries of the ME may depend upon the level of activity in the GnRH cells, the number of GnRH cells with access to the peripheral circulation will be examined under different levels of secretory activity. (2) Because synaptic connections relay environmental (and perhaps even steroidal) cues to GnRH neuroendocrine cells, we will continue our analysis of the synaptic input to the GnRH network. Specifically we will determine a) whether GnRH cells that project to ME receive a different pattern of innervation than those that do not, b) whether GnRH structures in the female are innervated more richly than in the male and c) what kinds of neurotransmitters impinge on GnRH cells. (3) GnRH neurons might also respond to appropriate signals at the cellular level by altering the processing of the GnRH precursor or at the genomic level by modulating transcription of mRNA. We shall employ specific antisera which recognize different amino acid sequences of the GnRH precursor to examine changes in cleavage patterns during different reproductive states. Ultrastructural immunocytochemical techniques will also be used to determine which sub-cellular compartments store and/or process the GnRH precursor protein. (4) To study transcription of the mRNA for GnRH we shall develop anti-mRNA probes (riboprobes) and quantitative in situ hybridization procedures. Throughout these experiments emphasis will be placed on a quantitative anatomical analysis for we consider it likely that different populations of GnRH neurons respond differentially to various physiological signals.